Beta-thalassemia is an inherited anemia in which synthesis of the hemoglobin beta-chain is decreased. Clinical features of beta-thalassemia include variably severe anemia and iron overload due to increased intestinal iron absorption, which may result in damage to vital organs. The hepatic peptide; hepcidin is a key regulator of iron metabolism in mammals. The present study aimed to determine the relationship between hepcidin expression and iron status in beta-thalassemia patients with hepatitis C virus infection. The study included 50 patients diagnosed as beta-thalassemia major (21 of them were HCV infected and 29 were HCV negative), in addition, 20 healthy subjects were enrolled in the study. The hepatic iron and hepcidin mRNA concentration in liver biopsy samples were measured, as well as serum ferritin, serum iron, hemoglobin and levels and serum hepcidin. Result showed remarkable decrease of serum and liver hepcidin mRNA expression in thalassemic patients as compared to controls, and showed a positive correlation with hemoglobin concentration, but negatively correlated with serum ferritin level and hepatic iron index (HII). In HCV infected patients, serum and liver hepcidin mRNA were markedly depressed in HCV positive beta-thalassemia cases, and positively correlated serum albumin and prothrombin concentrations, but inversely correlated with HII and fibrosis score. In HCV positive beta-thalassemia major patients, the hepcidin mRNA level was positively correlated with the synthetic function of the liver (namely serum albumin and prothrombin concentration) and with serum hepcidin level. While, both serum and hepcidin mRNA level was inversely correlated with HII and fibrosis score in these patients. These results suggest a possible role of hepcidin expression in iron overload in beta-thalassemia major, consequent disease progression and development of liver fibrosis.